The present invention relates to an NMR measurement method and apparatus for the same.
An NMR apparatus is an analyzer which irradiates predetermined atomic nucleus in a sample placed in a static magnetic field with a magnetic field pulse of a radio frequency and detects an NMR signal from the predetermined atomic nucleus after a predetermined time. In NMR spectrometry in recent years, multi-dimensional NMR measurement is widely conducted. The multi-dimensional NMR measurement displays an NMR signal in a frequency space having two or more frequency axes, and thereby improves resolution compared to one-dimensional NMR measurement, facilitates a spectral analysis and has an advantage of being able to elucidate interaction between atomic nuclear spins. This advantage is particularly important for NMR measurement targeted at a high polymer such as protein having a large molecular weight and complex spectrum.
FIG. 1 and FIG. 2 show schematic diagrams of 2-dimensional NMR measurement as an example of multi-dimensional NMR measurement. 3-dimensional or higher multi-dimensional measurement is an extension of 2-dimensional measurement and does not include any new concept, and therefore separate explanations will be omitted. FIG. 1 shows a general configuration of a pulse sequence (hereinafter referred to as “2-dimensional pulse sequence”) used for 2-dimensional NMR measurement. A two-dimensional pulse sequence consists of five periods; preparation period, evolution period, mixing period, acquisition period and relaxation period.
Of the five periods, the preparation period and mixing period each include irradiation of one or more magnetic field pulses. The evolution period is a delay time from the preparation period to the mixing period and normally expressed as t1 and called an “evolution period.” The acquisition period is a period during which an NMR signal is detected by the above described reception system and normally expressed as t2. Execution of a pulse sequence having one-time acquisition period is generally called “scan” and used as the unit for measuring the number of times NMR measurement is performed. The relaxation period is a waiting time (period) until the atomic nucleus returns to its original state before the irradiation of the pulse sequence.
2-dimensional NMR measurement is realized by repeating the two-dimensional pulse sequence while changing the evolution period t1. FIG. 2 is a schematic diagram thereof. Ns denotes a scan count, and the pulse sequence scan is repeated Na times with the evolution period t1 fixed. All the detected NMR signals are integrated using a reception processor. For this reason, Na is called an “integration count.” When Na scans are finished, the evolution period t1 is incremented by an increment Δt1 entered by the user beforehand and Na scans are performed again. 2-dimensional NMR measurement is realized by repeating this process until the evolution period t1 is incremented the number of times entered by the user beforehand, that is, Nt1 times.
As a result, 2-dimensional NMR measurement requires Na×Nt1 scans. Multi-dimensional NMR measurement generally includes (number of dimensions-1) evolution times and (number of dimensions-1) increment counts Ni's. For example, in the case of 3-dimensional NMR measurement, there are two evolution periods (times) of t1 and t3 and two increment counts of Nt1 and Nt3. The scan count necessary for 3-dimensional NMR measurement is Nscan=Na×Nt1×Nt3. When Δt1 is generalized and called “Δti” and Nt1 and Nt3 or the like are generalized and called “Nti”, the number of scans necessary for D-dimensional NMR measurement is on the order of NtiD−1.
While multi-dimensional NMR measurement including 2-dimensional NMR measurement has an advantage that resolution is improved and it is possible to elucidate interaction between atomic nuclear spins, it requires several tens to several thousands of scans to obtain a result, which takes a considerable time. The scan count necessary for multi-dimensional NMR measurement increases by increment count Nt1 to the (number of dimensions-1)th power, and therefore it is important to reduce Nti and the number of dimensions in multi-dimensional NMR measurement.
In order to reduce Nti, folding and aliasing of a signal and a half-dwell detection method are conventionally used. According to the conventional technique described in Ad Bax, Mitsuhiko Ikura, Lewis Kay, and Guang Zhu, Removal of F1 Baseline Distortion and Optimization of Folding in Multidimensional NMR Spectra, Journal of Magnetic Resonance, vol. 91, PP. 174-178 (1991), when Nti is reduced without deteriorating frequency resolution, the observable frequency bandwidth becomes narrower. This results in “folding and aliasing” of a signal on a frequency spectrum in which a signal appears at a frequency different from its original frequency. When folding and aliasing of a signal occurs, a frequency discrimination method for identifying the original frequency of the folded and aliased signal is necessary. The conventional technique uses the half-dwell detection method as the frequency discrimination method.
The half-dwell detection method conducts multi-dimensional NMR measurement with only the first time increment set to half the original increment Δti, that is, Δti/2. According to the half-dwell detection method, when the first acquisition time is changed, the sign of a spectrum whose folding and aliasing count is an odd number is inverted, whereas the sign of a spectrum whose folding and aliasing count is an even number remains unchanged.
Therefore, it is possible to distinguish the folded and aliased spectrum using this difference in the sign. In this way, the conventional technique can reduce Nti and the scan count necessary for multi-dimensional NMR measurement using the signal folding and aliasing and half-dwell detection method without deteriorating frequency resolution or without mistakenly discriminating the signal frequency. The relationship between signal folding and aliasing, Nti, frequency resolution and signal frequency discrimination is explained in John Cavanagh, Wayne J. Faribrother, Arthur G. Palmer III, and Nicholas J. Skelton, CHAPTER 4, Multidimensional NMR Spectroscopy, PP. 227-236 (Academic Press, 1995).
The conventional method using the signal folding and aliasing and half-dwell detection can distinguish whether a folding and aliasing count is an odd number or even number, but cannot distinguish the folding and aliasing count. Therefore, even under an optimal condition, the available folding and aliasing count is once at most. In the conventional technique using folding and aliasing, it is basically impossible to reduce Nti to less than ½ through predetermined NMR measurement alone without any problem with a spectral analysis.
When signals overlap due to folding and aliasing, there is another problem that it is difficult to decide signal signs. Overlapping of signals is more likely to occur as a spectrum becomes more complex, and therefore it is particularly difficult to decide folding and aliasing using signal signs particularly for high polymers having a complex spectrum. Avoiding this problem requires folding and aliasing to be reduced, and therefore it is actually difficult for the conventional technique to reduce Nti to close to 1/2.
The technique of deciding folding and aliasing using sign inversion of signals is effective for a conventional NMR apparatus having a low signal to noise ratio. Since Nscan needs to be increased to complement a low signal to noise ratio, it is a signal to noise ratio and not Nti of multi-dimensional measurement that constitutes a major factor in determining Nscan. For this reason, there has been no need to reduce Nti to close to 1/2 or below 1/2.
However, the signal to noise ratio of the NMR apparatus has been improved considerably and it is necessary to drastically reduce Nti compared to the conventional technique in order to improve the throughput of multi-dimensional NMR measurement. It is an object of the present invention to provide a measurement method and apparatus capable of reducing N to ½ with only predetermined NMR measurement without any problem with spectral analysis, and reduce a scanning count necessary for multi-dimensional NMR measurement and improve the throughput of multi-dimensional NMR measurement.